Early Alterations Of Microrna Expression To Predict And Modulate Melanoma Metastasis.

8550 Background: Melanoma is curable for most patients whose tumors are surgically removed early in disease progression; however, many primary melanomas recur and progress to metastasis. Clinical staging is insufficient to robustly classify patients at highest risk of recurrence, and prognostic molecular biomarkers have not yet been identified. Methods: We performed miRNA profiling of 92 FFPE primary melanomas to discover metastasis relevant miRNAs and develop predictive models of recurrence, which were then validated in an independent cohort. We identified miRNAs differentially expressed between primary tumors that did and did not recur (3-year minimum follow-up) and between thick and thin lesions. We selected candidate miRs for screening in a fluorescence-based in vitro invasion assay, and prioritized a subset for in vivo testing. Potential downstream mediators of these miRNAs were selected by mRNA array analysis and tested in a secondary invasion screen. mRNA candidates that mimicked the miR’s invasion-suppressive effect were tested in 3’UTR reporter assays to confirm them as direct targets. Results: Using the discovery cohort we identified a 20 miRNA signature that can distinguish Stage I/II primary tumors (n=70) that did from those did not recur with 3-year minimum follow-up with an AUC=94%, 95% CI: (0.88, 0.99). Applying this model to predict risk for recurrence in the independent validation cohort yielded an AUC = 96%, 95% CI: (0.90, 1) in discriminating recurrent versus non-recurrent stages I/ II patients (n=45). From the discovery cohort, 40 candidates were selected for invasion assay screening, of which 5 miRNAs robustly inhibit in vitro invasion in 5 melanoma cell lines. Three miRs (miR-382, miR-516b, and miR-7) strongly suppressed metastasis in a mouse model. Moreover, multiple mRNAs tested as potential mediators mimicked the invasion-suppressive effects of candidate miRs. Of those, NCAPG2 and CDC42 were identified as miR-516b targets, CTTN as a miR-382 target, and PIK3CD as a miR-7 target. These genes have been linked to metastasis in melanoma or other tumors. Conclusions: Our data demonstrate that aberrant expression of specific miRNAs at diagnosis is predictive of and functionally impacts melanoma progression.